Abstract Parkinson's disease (PD) is an important cause of morbidity in the Veteran population. Current treatments do not prevent disability resulting from progression of PD. Understanding the pathogenesis of the common sporadic form of PD is therefore an urgent objective, in order that appropriate therapeutic interventions can be developed. Multiple convergent lines of evidence suggest that impairment of mitochondrial respiratory chain function and abnormalities of -synuclein metabolism may be central to sporadic PD. Although evidence in vitro suggests the presence of bidirectional interactions between -synuclein and the mitochondrion, it is unclear whether these underlie dopaminergic neuronal degeneration in PD. In this proposal, we will exploit two recent technical advances in order to ask whether -synuclein is an important factor in the dopaminergic neuronal degeneration that occurs in vivo in response to partial systemic mitochondrial impairment, of the type found in sporadic PD. First, we have developed novel adeno-associated virus (AAV) vectors that express short hairpin RNA (shRNA) targeting the endogenous rat -synuclein transcript, and have optimized in vivo delivery of AAV vectors to the rat substantia nigra, allowing modulation of - synuclein expression in the rat nigro-striatal projection in vivo. Second, we have developed a novel model of sporadic PD, in which partial systemic mitochondrial complex I impairment, induced by daily intraperitoneal rotenone delivery, causes a highly reproducible lesion of the substantia nigra, including formation of prominent -synuclein immunoreactive inclusions resembling Lewy bodies and degeneration of dopaminergic neurons. Since it reproducibly recapitulates multiple relevant abnormalities characteristic of sporadic PD, the intraperitoneal rotenone model will be uniquely suitable for determination of whether -synuclein represents a critical mediator in the pathogenesis of the disease. In the proposed experiments, we will examine the behavioral, neurochemical and histological effects of -synuclein knockdown in an adult animal (objective 1). We anticipate from data in -synuclein knockout mice that there will be a subtle reduction in dopaminergic neurotransmission. However, developmental compensatory/adaptive changes that may be present in germline null mutants will be absent from these experiments in adults; consequently, it is possible that findings will be different to those reported in mouse knockouts. These data will form a basis for interpretation of subsequent experiments, and will inform on the likelihood of potential safety issues complicating neuroprotective strategies directed at modulation of -synuclein expression in the adult brain. We will next ask whether -synuclein knockdown is protective against the formation of -synuclein inclusions, neurobehavioral deficits and nigral cell loss after chronic rotenone exposure (objective 2). We hypothesize that pathogenesis of the nigral lesion is dependent on a key interaction between mitochondria and -synuclein, and consequently that reduction of endogenous -synuclein expression following shRNA vector transduction will ameliorate pathology, resulting in significant preservation of dopaminergic function. This experiment will inform on the possible efficacy of reducing cellular - synuclein levels or targeting abnormal functions of -synuclein in preventing PD progression. Together, these studies will determine the importance of an interaction in vivo between two major characteristic abnormalities of PD in the pathogenesis of neurodegeneration, and will determine whether -synuclein represents a valid therapeutic target for neuroprotection in the common sporadic form of Parkinson's disease.